Tauopathies are a group of neurodegenerative diseases that are characterized pathologically by neuronal loss and the accumulation of aggregates of the microtubule-associated protein Tau in surviving CNS neurons. Collectively, these disease, which include progressive supranuclear palsy (PSP), frontotemporal dementia (FTD), chronic traumatic encephalopathy (CTE) and Alzheimer's disease (AD), are a common and important cause of progressive neurological disability in Veterans. Current treatments only partially mitigate symptoms and do not alter the course of these diseases; consequently, there is an urgent need for new approaches that address pathogenic mechanisms and prevent disease progression. Convergent genetic and pathological evidence strongly suggests that Tau is centrally involved in causing neurodegeneration in these diseases, and there is accumulating evidence that oligomeric Tau species are a key pathogenic mediator. In addition, human tauopathy tissue and experimental Tau models both show striking evidence of impaired mitochondrial function resulting in reactive oxygen species (ROS) production and oxidative damage to neurons. However, there is no consensus regarding the mechanisms by which Tau and mitochondria interact and it is not known whether changes in mitochondrial function are a proximate pathogenic event that could be targeted therapeutically. Building on: (i) our recent finding that cytoplasmic protein oligomers inhibit mitochondrial protein import by interacting with the translocase of the outer mitochondrial membrane (TOMM) complex; (ii) published evidence showing that oligomeric Tau species are neurotoxic and impair expression of mitochondrial proteins; and (iii) our preliminary data showing that Tau interacts with the TOMM complex in Alzheimer's disease tissue, our overall guiding hypothesis is that: inhibition of mitochondrial protein import by oligomeric Tau causes progressive mitochondrial functional deficits that provoke oxidative damage and neuronal death. To address this hypothesis directly in vivo, we have generated novel zebrafish tauopathy models with progressive and highly reproducible phenotypes, allowing establishment of the sequence of biochemical events underlying pathogenesis over a practicable experimental time course. This model has been constructed on a genetic background that lacks pigment, allowing imaging of CNS neurons in vivo by confocal microscopy. Coupled with novel transgenic zebrafish lines expressing ratiometric reporters of cellular mitochondrial function, we will determine how neuronal mitochondrial bioenergetic functions, ROS production, and cellular RedOx status change during progressive tauopathy the CNS in vivo, and whether abnormal phenotypes can be rescued by over-expression of the translocase complex protein TOM20 (Aim 1). These experiments will test the hypothesis that protein import-dependent abnormalities of mitochondrial function in neurons precede the onset of neurological phenotypes in tauopathy and worsen with disease progression, causing oxidative damage and cell death. Next, in a series of complementary experiments in vitro and in vivo, we will test whether oligomeric Tau competes with nuclear-encoded mitochondrial proteins for interaction with TOM20 to directly inhibit mitochondrial protein import (Aim 2). Finally, exploiting the translational potential of Ta transgenic zebrafish, we will carry out the first phenotype-based drug re-purposing screen in a vertebrate tauopathy model (Aim 3). Using novel, automated neurological phenotyping assays we will interrogate a chemical library against functional endpoints that are caused by tauopathy, but which are unbiased to preconceptions regarding molecular mechanisms of pathogenesis. Compounds that are identified as modulators of tauopathy in vivo in this pilot screen will be tested for activity in restoring mitochondrial protein import and function, and will be candidates for further therapeutic development. Together, these studies will elucidate mitochondrial mechanisms underlying degeneration of CNS neurons in tauopathy in vivo and will provide new avenues of investigation for the development of effective therapeutics.